sequence. If the sequence was <25% GC it was assigned a score of 0.01 if it was <55% it was assigned a score of 3, if it was <60% it was assigned a score of 1, and if >60% it was assigned a score of 0.01. (iii) A “4-mer penalty” of 0.01 was assigned for any hairpin containing the same nucleotide in 4 subsequent nucleotides. (iv) A “7 GC penalty” of 0.01 was assigned to any hairpin containing any 7 consecutive G/C nucleotides. (v) We removed all hairpins containing an A in either position 1 or position 2 of the hairpin. (vi) We removed all hairpins containing a repeat masked nucleotide. (vii) Finally, we computed a “miRNA-seed penalty” by looking at the forward positions 11–17, 12–20, and 13–19 of the hairpin as well as the reverse complement of positions 14–20, 15–21, or 16–21 plus a 3′ C. We then looked up whether these positions matched known miRNA seeds and with what frequency. We computed the scores for the forward and reverse positions and defined the score as the product of the forward and reverse scores. The final score for each hairpin sequence is defined as the product of all seven
